A Microfluidic-Based Multi-Shear Device for Investigating the Effects of Low Fluid-Induced Stresses on Osteoblasts

Interstitial fluid flow (IFF) within the extracellular matrix (ECM) produces low magnitude shear stresses on cells. Fluid flow-induced stress (FSS) plays an important role during tissue morphogenesis. To investigate the effect of low FSS generated by IFF on cells, we developed a microfluidic-based cell culture device that can generate multiple low shear stresses. By changing the length and width of the flow-in channels, different continuous low level shear stresses could be generated in individual cell culture chambers. Numerical calculations demonstrate uniform shear stress distributions of the major cell culture area of each chamber. This calculation is further confirmed by the wall shear stress curves. The effects of low FSS on MC3T3-E1 proliferation and differentiation were studied using this device. It was found that FSS ranging from 1.5 to 52.6 µPa promoted MC3T3-E1 proliferation and differentiation, but FSS over 412 µPa inhibited the proliferation and differentiation of MC3T3-E1 cells. FSS ranging from 1.5 to 52.6 µPa also increased the expression of Runx2, a key transcription factor regulating osteoblast differentiation. It is suggested that Runx2 might be an important regulator in low FSS-induced MC3T3-E1 differentiation. This device allows for detailed study of the effect of low FSS on the behaviors of cells; thus, it would be a useful tool for analysis of the effects of IFF-induced shear stresses on cells.

[1]  M. Neeman,et al.  Overexpression of vascular endothelial growth factor 165 drives peritumor interstitial convection and induces lymphatic drain: magnetic resonance imaging, confocal microscopy, and histological tracking of triple-labeled albumin. , 2002, Cancer research.

[2]  L K Chin,et al.  Production of reactive oxygen species in endothelial cells under different pulsatile shear stresses and glucose concentrations. , 2011, Lab on a chip.

[3]  H J Donahue,et al.  Osteopontin Gene Regulation by Oscillatory Fluid Flow via Intracellular Calcium Mobilization and Activation of Mitogen-activated Protein Kinase in MC3T3–E1 Osteoblasts* , 2001, The Journal of Biological Chemistry.

[4]  Menahem Y. Rotenberg,et al.  A multi-shear perfusion bioreactor for investigating shear stress effects in endothelial cell constructs. , 2012, Lab on a chip.

[5]  B Lepioufle,et al.  Study of osteoblastic cells in a microfluidic environment. , 2006, Biomaterials.

[6]  S MacNeil,et al.  Matrix production and collagen structure are enhanced in two types of osteogenic progenitor cells by a simple fluid shear stress stimulus. , 2012, European cells & materials.

[7]  Fluid shear of low magnitude increases growth and expression of TGFbeta1 and adhesion molecules in human bone cells in vitro. , 2004, Experimental and clinical endocrinology & diabetes : official journal, German Society of Endocrinology [and] German Diabetes Association.

[8]  G. Whitesides.  The origins and the future of microfluidics , 2006, Nature.

[9]  Jinseok Heo,et al.  Spatially resolved shear distribution in microfluidic chip for studying force transduction mechanisms in cells. , 2010, Lab on a chip.

[10]  T M Keaveny,et al.  Osteoblasts respond to pulsatile fluid flow with short-term increases in PGE(2) but no change in mineralization. , 2001, Journal of applied physiology.

[11]  J A Frangos,et al.  Review: Bone tissue engineering: The role of interstitial fluid flow , 1994, Biotechnology and bioengineering.

[12]  Melody A. Swartz,et al.  Interstitial fluid flow induces myofibroblast differentiation and collagen alignment in vitro , 2005, Journal of Cell Science.

[13]  W. R. Thompson,et al.  Mechanical regulation of signaling pathways in bone. , 2012, Gene.

[14]  Kyung-Jin Jang,et al.  Fluid-shear-stress-induced translocation of aquaporin-2 and reorganization of actin cytoskeleton in renal tubular epithelial cells. , 2011, Integrative biology : quantitative biosciences from nano to macro.

[15]  G V Cochran,et al.  Interstitial fluid flow as a factor in decubitus ulcer formation. , 1981, Journal of biomechanics.

[16]  Yunyan Xie,et al.  Fluid flow stress induced contraction and re-spread of mesenchymal stem cells: a microfluidic study. , 2012, Integrative biology : quantitative biosciences from nano to macro.

[17]  Yong Woo Lee,et al.  Effect of intermittent shear stress on mechanotransductive signaling and osteoblastic differentiation of bone marrow stromal cells. , 2008, Tissue engineering. Part A.

[18]  G. Karsenty,et al.  Osf2/Cbfa1: A Transcriptional Activator of Osteoblast Differentiation , 1997, Cell.

[19]  Makoto Sato,et al.  Targeted Disruption of Cbfa1 Results in a Complete Lack of Bone Formation owing to Maturational Arrest of Osteoblasts , 1997, Cell.

[20]  Axel Rosenhahn,et al.  Quantification of the adhesion strength of fibroblast cells on ethylene glycol terminated self-assembled monolayers by a microfluidic shear force assay. , 2010, Physical chemistry chemical physics : PCCP.

[21]  Hui Zhao,et al.  Integrated microfluidic chip for endothelial cells culture and analysis exposed to a pulsatile and oscillatory shear stress. , 2009, Lab on a chip.

[22]  Umut A. Gurkan,et al.  The Mechanical Environment of Bone Marrow: A Review , 2008, Annals of Biomedical Engineering.

[23]  Antonios G Mikos,et al.  In vitro generated extracellular matrix and fluid shear stress synergistically enhance 3D osteoblastic differentiation. , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[24]  T. Takano-Yamamoto,et al.  Fluid Shear Stress Induces Less Calcium Response in a Single Primary Osteocyte Than in a Single Osteoblast: Implication of Different Focal Adhesion Formation , 2006, Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research.

[25]  R K Jain,et al.  Direct measurement of interstitial convection and diffusion of albumin in normal and neoplastic tissues by fluorescence photobleaching. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[26]  K. Jensen,et al.  Cells on chips , 2006, Nature.

[27]  Elise F. Morgan,et al.  Assessment of a mechano-regulation theory of skeletal tissue differentiation in an in vivo model of mechanically induced cartilage formation , 2009, Biomechanics and modeling in mechanobiology.

[28]  Jingjun Xu,et al.  A multishear microfluidic device for quantitative analysis of calcium dynamics in osteoblasts. , 2011, Biochemical and biophysical research communications.

[29]  Joong Yull Park,et al.  Cell morphological response to low shear stress in a two-dimensional culture microsystem with magnitudes comparable to interstitial shear stress. , 2010, Biorheology.

[30]  Tal Dvir,et al.  Activation of the ERK1/2 cascade via pulsatile interstitial fluid flow promotes cardiac tissue assembly. , 2007, Tissue engineering.

[31]  G. Reilly,et al.  The effects of mechanical loading on mesenchymal stem cell differentiation and matrix production. , 2011, Vitamins and hormones.

[32]  Jason W. Triplett,et al.  Osteoblasts and osteocytes respond differently to oscillatory and unidirectional fluid flow profiles , 2007, Journal of cellular biochemistry.

[33]  R. Bareille,et al.  Responsiveness of human bone marrow stromal cells to shear stress , 2009, Journal of tissue engineering and regenerative medicine.

[34]  A. Papadimitropoulos,et al.  Effects of fluid flow and calcium phosphate coating on human bone marrow stromal cells cultured in a defined 2D model system. , 2008, Journal of biomedical materials research. Part A.

[35]  Christopher R Jacobs,et al.  Mechanically induced osteogenic differentiation – the role of RhoA, ROCKII and cytoskeletal dynamics , 2009, Journal of Cell Science.

[36]  Douglas Lauffenburger,et al.  Interstitial fluid flow intensity modulates endothelial sprouting in restricted Src-activated cell clusters during capillary morphogenesis. , 2009, Tissue engineering. Part A.

[37]  Benoît Carpentier,et al.  Bioreactors for Bone Tissue Engineering , 2011, The International journal of artificial organs.

[38]  R. Kwon,et al.  Microfluidic Enhancement of Intramedullary Pressure Increases Interstitial Fluid Flow and Inhibits Bone Loss in Hindlimb Suspended Mice , 2010, Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research.